Previous data from our laboratory indicate that nitric oxide (NO) produced in the renal medulla plays a critical role in the regulation of fluid and electrolyte homeostasis and the long term control of arterial pressure. However, the relative contribution of the three constitutively expressed isoforms of NOS to the control of renal tubular and vascular function in this region of the kidney is unknown. This project will address the hypothesis that the three isoforms of NOS are localized in different cell types within the renal medulla, and that the NO produced by these isoforms serves as an autocrine controller of tubular or vascular function to regulate sodium excretion and consequently blood pressure. A corollary of this hypothesis is that selective inhibition of different NOS isoforms should alter tubular transport or medullary blood flow depending on where a particular isoform is expressed. This hypothesis will be addressed using a unique, integrative, biological approach. The initial set of experiments will determine the dose of each of the NOS isoform inhibitors which when infused into the renal medullary interstitium selectively inhibits the targeted isoform and elicits effects on sodium excretion and medullary blood flow. We will then utilize chronic renal medullary interstitial infusion of these agents to determine the role of each of the NOS isoforms (nNOS, iNOS and eNOS) in influencing "salt-sensitivity" of blood pressure, and their role in the long term control of sodium balance and arterial pressure in conscious rats. The chronic functional effects of isoform- selective inhibition will be correlated with changes in renal medullary NO concentration measured in conscious rats using a new microdialysis technique and the expression of NOS proteins and enzyme activity in the medulla. Finally the mechanisms (either tubular or vascular) by which chronic inhibition of various NOS isoforms in the renal medulla leads to sodium retention and hypertension will be determined by examining the pressure-natriuresis relationship, tubular function using micropuncture, medullary blood flow using laser Doppler flowmetry and renal interstitial hydrostatic pressure (RIHP) using implanted capsules. This unique array of biochemical, molecular and integrative biological techniques should provide important new information regarding the distribution of various isoforms of NOS in the renal medulla and the critical importance of these isoforms in the regulation of medullary blood flow, tubular function and the long term control of arterial pressure.